Spinal muscular atrophy (SMA) is a common and often fatal, motor neuron degenerative disease and a leading genetic cause of infant mortality (Talbot et al., 2001, Semin Neurol 21(2):189-97; Wirth et al., 2006, Semin Pediatr Neurol 13(2):121-31). SMA severity corresponds to the degree of functional survival of motor neurons (SMN) protein deficiency. SMN is a ubiquitously expressed protein that plays a critical role in RNA metabolism and is essential for viability of all cells in eukaryotes (Neuenkirchen et al., 2008, FEBS Lett 582(14):1997-2003; Yong, et al., 2004, Trends Cell Biol 14(5):226-32). As part of a large multi-protein complex, the SMN complex, SMN functions in the biogenesis of snRNPs, the major subunits of the spliceosome (Meister et al., 2001, Nat Cell Biol 3(11):945-9; Pellizzoni et al., 2002, Science 298(5599):1775-9; Liu et al., 1997, Cell 90(6):1013-21; Fischer et al., 1997, Cell 90(6):1023-9).
Although SMN deficiency manifests itself as a motor neuron disease, its molecular consequences are evident as profound disruptions in RNA metabolism in all tissues tested in a SMA mouse model (Zhang et al., 2008, Cell 133(4):585-600). There are two SMN genes in humans, SMN1 and SMN2, both encoding the same open reading frame. The vast majority of SMA patients have homozygous SMN1 deletions and are sustained by one or more copies of SMN2. However, due to a C/T substitution at position 6 of exon 7 that does not change the encoded amino acid, the splicing of the SMN2 pre-mRNA incurs frequent (−80%) exon 7 skipping. This produces an SMN protein (SMNΔ7) that lacks the normal carboxyl-terminal 16 amino acids, and acquires instead four amino acids, EMLA, encoded by exon 8 (Le et al., 2005, Hum Mol Genet 14(6):845-57). Thus, BIM deletions expose the splicing defect of SMN2 and its ineffectiveness in producing full-length normal SMN protein (Wirth et al., 2006, Semin Pediatr Neurol 13(2):121-31; Cooper et al., 2009, Cell 136(4):777-93).
Biochemical experiments in vitro suggest that SMNΔ7 is not fully functional compared to normal SMN protein, including a diminished oligomerization and binding to protein substrates, such as the snRNP Sm proteins (Pellizzoni et al., 1999, Proc Natl Acad Sci USA 96(20):11167-72; Lorson et al., 1998, Nat Genet 19(1):63-6).
Increased SMN2 copy number correlates with a milder clinical phenotype in SMA patients (Wirth et al., 2006, Hum Genet 119(4):422-8). Furthermore, studies in cells suggest (Wang et al., 2001, J Biol Chem 276(48):45387-93) and experiments in SMN-deficient mice demonstrate that expression of an increasing copy number of SMNΔ7 cDNA transgenes proportionately lessens SMA severity (Le et al., 2005, Hum Mol Genet 14(6):845-57). This suggests that even a modest SMNΔ7 increase is beneficial in SMA.
There is a long-standing need to identify the cause of SMNΔ7 instability in order to correct the deficit in vivo. The present invention fills this need.